Sterols (STs) have a key role in regulating the fluidity and permeability of membranes in plants (phytosterols) that have wide structural diversity. We studied the effect of structural STs diversity on salt tolerance in halophytes. Specifically, we used gas chromatography–mass spectrometry (GC–MS), including two-dimensional gas chromatography–mass spectrometry (GCxGC–MS), to assess the STs composition in leaves of 21 species of wild-growing halophytes from four families (Asteraceae, Chenopodiaceae, Plumbaginaceae, Tamaricaceae) and three ecological groups (Euhalophytes (Eu), recretophytes (Re), salt excluders (Ex)). Fifteen molecular species of STs from three main groups, Δ⁵-, Δ⁷-and Δ⁰- STs (stanols), were detected. Plants of the genus Artemisia were characterized by a high content of stigmasterol (30–49% of the total STs), while β-sitosterol was the major compound in two Limonium spp., where it comprised 84–92% of the total STs. Species of Chenopodiaceae were able to accumulate both Δ⁵-and Δ⁷-STs and stanols. The content of the predominant Δ⁵-STs decreased in the order Ex → Re → Eu. Molecular species with a saturated steroid nucleus were identified in Eu and Re, suggesting their special salt-accumulating and salt-releasing functions. The structural analogues of stigmasterol, having a double bond C-22, were stigmasta-7,22-dien-3β-ol (spinasterol) and stigmast-22-en-3β-ol (Δ^7--sitosterol). The ratio of Δ⁵-stigmasterol/Δ⁵-β-sitosterol increased in Ex plants, and spinasterol/Δ^7--sitosterol and 22-stigmastenol/sitostanol increased in Eu plants. These data support the well-known role of stigmasterol and its isomers in plant responses to abiotic and biotic factors. The variability in STs types and their ratios suggested some involvement of the sterol membrane components in plant adaptation to growth conditions. The balance of Δ⁵-, Δ⁷-and stanols, as well as the accumulation of molecular analogues of stigmasterol, was suggested to be associated with salt tolerance of the plant species in this investigation.